PO.TB10.15 · 肿瘤生物学
Pyk2 通过囊泡运输在胶质母细胞瘤细胞中调控 Rab 蛋白
Pyk2 controls rab proteins by vesicle trafficking in glioblastoma cells
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
胶质母细胞瘤（GBM）是最具侵袭性的原发性脑肿瘤，中位生存期不足一年。GBM 的进展依赖于肿瘤细胞与其微环境之间持续的通讯，主要通过细胞外囊泡（EV）介导。这些囊泡通常含有 CCL2、CCL12 和 CCL5 等趋化因子，可吸引免疫细胞。这些免疫细胞一旦被募集，即可被肿瘤来源的信号重编程以支持肿瘤生长。Pyk2 通过与 Rab GTP 酶（细胞内囊泡转运的调控因子）相互作用来调控 EV 的运输和释放。通过激活 RhoA 和重组肌动蛋白，Pyk2 促进囊泡向质膜移动及其随后的分泌。我们假设 Pyk2 通过协调 Rab GTP 酶活性和肌动蛋白细胞骨架重塑来调控 GBM 细胞中的 EV 运输和分泌。我们分析了有和无 CRISPR/Cas9 介导的 Pyk2 敲除（Pyk2KO）的原代人 GBM 细胞系。分离 EV 并通过流式细胞术进行表征，使用 Avutometinib（1 µM）抑制 MEK/ERK 信号传导。共聚焦免疫荧光显示，Pyk2 促进 Rab27a 与 CD63⁺ 囊泡的结合，这与其在调控多囊泡体对接和外泌体释放中的作用一致。Pyk2 调控 Rab27a 的肌动蛋白依赖性运输，从而控制囊泡的转运和分泌。定量分析显示，Pyk2KO 使 EV 群体向更大的囊泡转变。Pyk2KO 降低了 EV 货物中 CCL2、CCL5、TNF 和 VEGF（TAM 激活和血管生成的介导因子）的水平。这些数据表明，Pyk2 调控 Rab27a 依赖性的内体囊泡对接和释放，而 MEK/ERK 信号传导则增强质膜来源的 EV 脱落。这些通路共同塑造了 EV 的大小和免疫调节货物。
Pyk2 通过囊泡运输在胶质母细胞瘤细胞中调控 Rab 蛋白。
查看英文原文 English abstract
Glioblastoma (GBM) is the most aggressive primary brain tumor, with a median survival of less than one year. GBM progression depends on constant communication between tumor cells and their microenvironment, primarily mediated through extracellular vesicles (EVs). These vesicles often contain chemokines such as CCL2, CCL12, and CCL5, which attract immune cells. Once recruited, these immune cells can be reprogrammed by tumor-derived signals to support tumor growth. Pyk2 regulates the trafficking and release of EVs by interacting with Rab GTPases, regulators of intracellular vesicle transport. Through activation of RhoA and reorganization of actin, Pyk2 facilitates vesicle movement toward the plasma membrane and their subsequent secretion. We hypothesized that Pyk2 regulates EV trafficking and secretion in GBM cells by coordinating Rab GTPase activity and actin cytoskeletal remodelling.Primary human GBM cell lines with and without CRISPR/Cas9-mediated Pyk2 knockout (Pyk2KO) were analysed. EVs were isolated and characterized by flow cytometry, MEK/ERK signaling was inhibited using Avutometinib(1µM). Confocal immunofluorescent revealed that Pyk2 promotes Rab27a association with CD63⁺ vesicles, consistent with its role in regulating multivesicular body docking and exosome release. Pyk2 regulates actin dependent trafficking of Rab27a thereby controlling vesicle transport and secretion. Quantitative analysis showed that Pyk2KO shifted the EV population toward larger vesicles. Pyk2KO reduced EV cargo levels of CCL2, CCL5, TNF, and VEGF, mediators of TAM activation and angiogenesis. These data indicate that Pyk2 regulates Rab27a-dependent endosomal vesicle docking and release, whereas MEK/ERK signaling enhances plasma membrane-derived EV shedding. Together, these pathways shape the size and immune regulatory cargo of EVs.
Pyk2 controls Rab proteins by vesicle trafficking in glioblastoma cells
利益披露 Disclosure
N. M. Ramirez Serrano, None..
L. Kucheryavykh, None.